Design,synthesis, biological evaluation,QSAR analysis and molecular modelling of new thiazol-benzimidazoles as EGFR inhibitors

Heterocyclic rings such as thiazole and benzimidazole are considered as privileged structures, since they constitute several FDA-approved drugs for cancer treatment. In this work, a new set of 2-(2-(substituted) hydrazinyl)-4-(1-methyl-1H-benzo[d]imidazol-2-yl) thiazoles 4a-q were designed as epidermal growth factor receptor (EGFR) inhibitors and synthesized using concise synthetic methods. The new target compounds have been evaluated in vitro for their suppression activity against EGFR TK. Compounds 4n, 4h, 4i, 4a and 4d exhibited significant potency in comparison with erlotinib which served as a reference drug (IC50, 71.67–152.59 nM; IC50 erlotinib, 152.59 nM). Furthermore, MTT assay revealed that compounds 4j, 4a, 4f, 4h, 4n produced the most promising cytotoxic potency against the human breast cancer cell line (MCF-7) (IC50; 5.96–11.91 µM; IC50 erlotinib; 4.15 µM). Compound 4a showed promising activity as EGFR TK inhibitor as well as anti-breast cancer agent. In addition, 4a induced apoptotic effect and cell cycle arrest at G2/M phase preventing the mitotic cycle in MCF-7 cells. Moreover, 4a upregulated the oncogenic parameters; caspase-3, p53, Bax/Bcl-2 as well as it inhibited the level of PARP-1 enzyme. QSAR study was carried out for the new derivatives and it revealed the goodness of the models. Furthermore, molecular docking studies represented the binding modes of the promising compounds in the active pocket of EGFR.     

In silico studies on p21-activated kinase 4 inhibitors: comprehensive application of 3D-QSAR analysis,molecular docking,molecular dynamics simulations,and MM-GBSA calculation

Abstract

P21-activated kinase 4 (PAK4) is a serine/threonine protein kinase, which is associated with many cancer diseases, and thus being considered as a potential drug target. In this study, three-dimensional quantitative structure-activity relationship (3D-QSAR), molecular docking and molecular dynamics (MD) simulations were performed to explore the structure-activity relationship of a series of pyrropyrazole PAK4 inhibitors. The statistical parameters of comparative molecular field analysis (CoMFA, Q ² = 0.837, R ² = 0.990, and R ² _pred = 0.967) and comparative molecular similarity indices analysis (CoMSIA, Q ² = 0.720, R ² = 0.972, and R ² _pred = 0.946) were obtained from 3D-QSAR model, which exhibited good predictive ability and significant statistical reliability. The binding mode of PAK4 with its inhibitors was obtained through molecular docking study, which indicated that the residues of GLU396, LEU398, LYS350, and ASP458 were important for activity. Molecular mechanics generalized born surface area (MM-GBSA) method was performed to calculate the binding free energy, which indicated that the coulomb, lipophilic and van der Waals (vdW) interactions made major contributions to the binding affinity. Furthermore, through 100?ns MD simulations, we obtained the key amino acid residues and the types of interactions they participated in. Based on the constructed 3D-QSAR model, some novel pyrropyrazole derivatives targeting PAK4 were designed with improved predicted activities. Pharmacokinetic and toxicity predictions of the designed PAK4 inhibitors were obtained by the pkCSM, indicating these compounds had better absorption, distribution, metabolism, excretion and toxicity (ADMET) properties. Above research provided a valuable insight for developing novel and effective pyrropyrazole compounds targeting PAK4.     

Synthesis and biological evaluation of 3-aryl-4-indolyl-maleimides as potent mutant isocitrate dehydrogenase-1 inhibitors

A series of 3-aryl-4-indolylmaleimide IDH1/R132H inhibitors with a novel structure was obtained by high-throughput screening and structure-based optimization. Most compounds such as 7a, 7d, 7h, 7i, 7k and 7o showed high inhibitory effects on IDH1/R132H and were highly selective against IDH1/WT, IDH2/WT, GDH, GK, and FBP. Evaluation of the biological activities and function at cellular level showed that compounds 7h, 7i and 7k could effectively suppress the production of 2-hydroxyglutaric acid in U87MG cells expressing IDH1/R132H. Additionally, 7h could reversed the differentiation block of the myeloid leukemic cell line, TF-1, caused by the overexpression of IDH1/R132H. We also explore the structure-activity relationship based on the experimental data, with an attempt to pave the way for future studies.     

Structure-activity relationships of 4-hydroxy-4-biaryl-proline acylsulfonamide tripeptides: A series of potent NS3 protease inhibitors for the treatment of hepatitis C virus

The design and synthesis of a series of tripeptide acylsulfonamides as potent inhibitors of the HCV NS3/4A serine protease is described. These analogues house a C4 aryl, C4 hydroxy-proline at the S2 position of the tripeptide scaffold. Information relating to structure-activity relationships as well as the pharmacokinetic and cardiovascular profiles of these analogues is provided.     

Phosphodiesterase inhibitors. Part 2: Design, synthesis, and structure-activity relationships of dual PDE3/4-inhibitory pyrazolo[1,5-a]pyridines with anti-inflammatory and bronchodilatory activity

A structural survey of pyrazolopyridine-pyridazinone phosphodiesterase (PDE) inhibitors was made with a view to optimization of their dual PDE3/4-inhibitory activity for respiratory disease applications. These studies identified (−)-6-(7-methoxy-2-trifluoromethylpyrazolo[1,5-a]pyridine-4-yl)-5-methyl-4,5-dihydro-3-(2H)-pyridazinone (KCA-1490, compound 2ac) as a compound with potent combined bronchodilatory and anti-inflammatory activity and an improved therapeutic window over roflumilast.     

Discovery of novel HCV inhibitors: Synthesis and biological activity of 6-(indol-2-yl)pyridine-3-sulfonamides targeting hepatitis C virus NS4B

A novel series of 6-(indol-2-yl)pyridine-3-sulfonamides was prepared and evaluated for their ability to inhibit HCV RNA replication in the HCV replicon cell culture assay. Preliminary optimization of this series furnished compounds with low nanomolar potency against the HCV genotype 1b replicon. Among these, compound 8c has identified as a potent HCV replicon inhibitor (EC₅₀ = 4 nM) with a selectivity index with respect to cellular GAPDH of more than 2500. Further, compound 8c had a good pharmacokinetic profile in rats with an IV half-life of 6 h and oral bioavailability (F) of 62%. Selection of HCV replicon resistance identified an amino acid substitution in HCV NS4B that confers resistance to these compounds. These compounds hold promise as a new chemotype with anti-HCV activity mediated through an underexploited viral target.     

Design,synthesis, and anti-cancer evaluation of new pyrido[2,3-d]pyrimidin-4(3H)-one derivatives as potential EGFRWT and EGFRT790M inhibitors and apoptosis inducers

A new series of pyrido[2,3-d]pyrimidin-4(3H)-one derivatives having the essential pharmacophoric features of EGFR inhibitors has been designed and synthesised. Cell viability screening was performed for these compounds against A-549, PC-3, HCT-116, and MCF-7 cell lines at a dose of 100 μM. The highest active derivatives (8a, 8 b, 8d, 9a, and 12b) were selected for IC₅₀ screening. Compounds 8a, 8 b, and 9a showed the highest cytotoxic activities and were further investigated for wild EGFR^WT and mutant EGFR^T790M inhibitory activities. Compound 8a showed the highest inhibitory activities against EGFR^WT and EGFR^T790M with IC₅₀ values of 0.099 and 0.123 µM, respectively. In addition, it arrested the cell cycle at pre-G1 phase and induced a significant apoptotic effect in PC-3 cells. Furthermore, compound 8a induced a 5.3-fold increase in the level of caspase-3 in PC-3 cells. Finally, docking studies were carried out to examine the binding mode of the synthesised compounds against both EGFR^WT and EGFR^T790M.     

Recognition of ribonuclease A by 3'-5'-pyrophosphate-linked dinucleotide inhibitors: a molecular dynamics/continuum electrostatics analysis

The proteins of the pancreatic ribonuclease A (RNase A) family catalyze the cleavage of the RNA polymer chain. The development of RNase inhibitors is of significant interest, as some of these compounds may have a therapeutic effect in pathological conditions associated with these proteins. The most potent low molecular weight inhibitor of RNase reported to date is the compound 5′-phospho-2′-deoxyuridine-3-pyrophosphate (P→5)-adenosine-3-phosphate (pdUppA-3′-p). The 3′,5′-pyrophosphate group of this compound increases its affinity and introduces structural features which seem to be unique in pyrophosphate-containing ligands bound to RNase A, such as the adoption of a syn conformation by the adenosine base at RNase subsite B₂ and the placement of the 5′-β-phosphate of the adenylate (instead of the α-phosphate) at subsite P₁ where the phosphodiester bond cleavage occurs. In this work, we study by multi-ns molecular dynamics simulations the structural properties of RNase A complexes with the ligand pdUppA-3′-p and the related weaker inhibitor dUppA, which lacks the 3′ and 5′ terminal phosphate groups of pdUppA-3′-p. The simulations show that the adenylate 5′-β-phosphate binding position and the adenosine syn orientation constitute robust structural features in both complexes, stabilized by persistent interactions with specific active-site residues of subsites P₁ and B₂. The simulation structures are used in conjunction with a continuum-electrostatics (Poisson-Boltzmann) model, to evaluate the relative binding affinity of the two complexes. The computed relative affinity of pdUppA-3′-p varies between −7.9 kcal/mol and −2.8 kcal/mol for a range of protein/ligand dielectric constants (ε_p) 2–20, in good agreement with the experimental value (−3.6 kcal/mol); the agreement becomes exact with ε_p = 8. The success of the continuum-electrostatics model suggests that the differences in affinity of the two ligands originate mainly from electrostatic interactions. A residue decomposition of the electrostatic free energies shows that the terminal phosphate groups of pdUppA-3′-p make increased interactions with residues Lys⁷ and Lys⁶⁶ of the more remote sites P₂ and P₀, and His¹¹⁹ of site P₁.     

Stereoisomeric flavour compounds LXVIII. 2-, 3-, and 4-Alkyl-branched acids,part 2: Chirospecific analysis and sensory evaluation

Enantioselective GC analysis of 4-ethyloctanoic and 4-methylheptanoic acid, using heptakis(2,3-di-O-methyl-6-O-tert-butyldimethylsilyl)-β-cyclodextrin as the chiral stationary phase, is described and the sensory properties of several 4-alkyl-branched acids, using gas chromatography-olfactometry (GC-O) equipment and octakis(2,3-di-O-methyl-6-tert-butyldimethylsilyl)-γ-cyclodextrin as the stationary phase, are evaluated. The chirospecific analysis of various 2-, 3-, and 4-alkyl-branched acids from commercially available Roman chamomile (Chamaemelum nobile (L.) Allioni), Parmesan cheese, and subcutaneous mutton adipose tissue, using either GC-GC (MDGC) or GC-MS analytical methods, is described. © 1994 Wiley-Liss, Inc.     

Responses of wild C4 and C3 grass (Poaceae) species to elevated atmospheric CO2 concentration: a meta-analytic test of current theories and perceptions

C4 plants contribute ≈ 20% of global gross primary productivity, and uncertainties regarding their responses to rising atmospheric CO₂ concentrations may limit predictions of future global change impacts on C4-dominated ecosystems. These uncertainties have not yet been considered rigorously due to expectations of C4 low responsiveness based on photosynthetic theory and early experiments. We carried out a literature review (1980–97) and meta-analysis in order to identify emerging patterns of C4 grass responses to elevated CO₂, as compared with those of C3 grasses. The focus was on nondomesticated Poaceae alone, to the exclusion of C4 dicotyledonous and C4 crop species. This provides a clear test, controlled for genotypic variability at family level, of differences between the CO₂-responsiveness of these functional types. Eleven responses were considered, ranging from physiological behaviour at the leaf level to carbon allocation patterns at the whole plant level. Results were also assessed in the context of environmental stress conditions (light, temperature, water and nutrient stress), and experimental growing conditions (pot size, experimental duration and fumigation method). Both C4 and C3 species increased total biomass significantly in elevated CO₂, by 33% and 44%, respectively. Differing tendencies between types in shoot structural response were revealed: C3 species showed a greater increase in tillering, whereas C4 species showed a greater increase in leaf area in elevated CO₂. At the leaf level, significant stomatal closure and increased leaf water use efficiency were confirmed in both types, and higher carbon assimilation rates were found in both C3 and C4 species (33% and 25%, respectively). Environmental stress did not alter the C4 CO₂-response, except for the loss of a significant positive CO₂-response for above-ground biomass and leaf area under water stress. In C3 species, stimulation of carbon assimilation rate was reduced by stress (overall), and nutrient stress tended to reduce the mean biomass response to elevated CO₂. Leaf carbohydrate status increased and leaf nitrogen concentration decreased significantly in elevated CO₂ only in C3 species. We conclude that the relative responses of the C4 and C3 photosynthetic types to elevated CO₂ concur only to some extent with expectations based on photosynthetic theory. The significant positive responses of C4 grass species at both the leaf and the whole plant level demand a re-evaluation of the assumption of low responsiveness in C4 plants at both levels, and not only with regard to water relations. The combined shoot structural and water use efficiency responses of these functional types will have consequential implications for the water balance of important catchments and range-lands throughout the world, especially in semiarid subtropical and temperate regions. It may be premature to predict that C4 grass species will lose their competitive advantage over C3 grass species in elevated CO₂.     

Detection and identification of 4‐hydroxy‐2‐nonenal Schiff‐base adducts along with products of Michael addition using data‐dependent neutral loss‐driven MS3 acquisition: Method evaluation through an in vitro study on cytochrome c oxidase modifications

We report a data‐dependent neutral‐loss‐driven MS³ acquisition to enhance, in addition to abundant Michael adducts, the detection of Schiff‐base adducts of proteins and 4‐hydroxy‐2‐nonenal, a reactive end product of lipid peroxidation. In vitro modification of cytochrome c oxidase, a mitochondrial protein complex, was used as a model to evaluate the method. The technique allowed for a confident validation of modification sites and also identified a Schiff‐base adduct in subunit Vb of the protein complex.     

Expression,SNV identification,linkage disequilibrium,and combined genotype association analysis of the muscle-specific gene CSRP3 in Chinese cattle

The cysteine and glycine-rich protein 3 (CSRP3) plays an important role in the myofiber differentiation. Here, we identified five SNVs in all exon and intron regions of the CSRP3 gene using DNA sequencing, PCR-RFLP and forced-PCR-RFLP methods in 554 cattle. Four of the five SNVs were significantly associated with growth performance and carcass traits of the cattle. In addition, we evaluated haplotype frequency and linkage disequilibrium coefficient of five sequence variants. The result of haplotype analysis demonstrated 28 haplotypes present in Qinchuan and two haplotypes in Chinese Holstein. Only haplotypes 1 and 8 were being shared by two populations, haplotype 14 had the highest haplotype frequency in Qinchuan (17.4%) and haplotype 8 had the highest haplotype frequency in Chinese Holstein (94.4%). Statistical analyses of combined genotypes indicated that some combined genotypes were significantly or highly significantly associated with growth and carcass traits in the Qinchuan cattle population. qPCR analyses also showed that bovine CSRP3 gene was exclusively expressed in longissimus dorsi muscle and heart tissues. The data support the high potential of the CSRP3 as a marker gene for the improvement of growth performance and carcass traits in selection programs.